Neurotropic growth factors such as nerve growth factor (NGF) and neutrophins stimulate proneuritic signals of neural cells (NCs) via ligand-induced dimerizations of neural specific membrane receptor- tyrosine kinases (nsmRTKs) and modulate several key processes in neural cells including axon specification and elongation. However, it is still unclear how these biomolecular regulators coordinate highly complex morphogenesis for the neural circuit network formation. To understand molecular mechanisms of axonal development with high spatio-temporal complexity, it is required to monitor neural cells in real-time at the single molecule level with multivariate analytical capability. Here, we propos an innovative approach to investigate molecular mechanisms of axonal development at the single molecule level using in situ surface plasmon dark field microscopy. Unlike conventional microscopy that only provides a static assessment of cell status, the proposed surface plasmon microscopy allows us to visualize single molecule events of the signaling molecules continuously over the axon generation period as well as morphogenetic development. With this new technique, specifically, I will challenge the following topics: 1. In situ monitoring of TrKA dimerization of individual nerual cells and its correlation with distinct morphogenesis (axon specification and elongation). 2. Focused neurite activation of a single cell and its communication with surrounding cells. 3. Test the feasibility of an artificial magnetic tweezer system for the guided axonal development. PUBLIC HEALTH RELEVANCE: The proposed optical probes with extreme brightness and photostability will provide specific and sensitive diagnostic tools for membrane protein dimerization. Moreover, the single-molecule studies of axon specification will may facilitate the design of efficient neurdegenerative drugs with minimal off-target effects for therapeutic use.